This invention relates to method and means for aiding the separation of blood components in a blood bag during centrifugation.
Human blood is separated into its various components in order to maximize the benefits of this valuable resource and to provide only the component required by an individual patient. For example, whole blood is typically processed into platelets, plasma and red blood cells.
The cellular components of blood have different densities. With the aid of a centrifuge, the various cell types establish themselves in layers according to their densities. Predetermined and well known centrifuge speed and time ratios are used to accomplish this separation. The red blood cells (RBC), being the most dense of the blood components, settle to the bottom of the fluid column. Above the RBC layer is formed the socalled "buffy coat" and the plasma layer forms above the buffy coat. If the correct time is used in conformance with the normal procedure for separating blood components, platelets are suspended in the plasma layer. After the first spin, the platelet-rich plasma (PRP) is transferred to an empty satellite bag and is centrifuged again at a higher speed for a longer period to further separate the platelet rich plasma (PRP) into platelet-poor plasma (PPP) and platelet concentrate (PC).
In order to harvest the maximum number of platelets from the PRP, the PRP is spun to produce a centrifugal force ranging from 3000 G to 4400 G for eight minutes in the first instance to five minutes in the second instance. This time/speed ratio almost always insures adequate platelet yield, but it results in the platelets impacting one another and clumping together, forming what is commonly called a platelet "button" at the bottom of the primary bag. The degree of centrifugation and resulting severity of button formation determine the degree of platelet damage. The higher the speed and the longer the time the platelets are subjected to that force, the higher the level of loss of platelet viability. Numerous researchers have recommended devising methods of reducing the cell damage caused by the such known harvesting methods, but no workable method has been developed heretofore.
Red blood cells can be separated still further. More particularly, after the separation of PRP, the residual RBC mass is comprised of approximately 70% RBC and 30% plasma. This mass may be centrifuged again at a higher speed, greater than 4000 G, for fifteen to thirty minutes to further separate them according to their agedependent density. The youngest (least dense) RBC's are called neocytes and migrate to the top of the RBC column while the oldest (most dense RBC's), called gerocytes, migrate to the bottom of the RBC column. The residual plasma found in the the RBC mass prior to this last spin is found above the young RBC mass. The older RBC mass is almost totally devoid of plasma.
The benefits of using only young RBCs in the treatment of certain disorders is known. The red blood cells or erythrocytes in donor blood have a certain life span. Actually, human blood contains more or less equal portions of red blood cells of ages between about 0 and 120 days. Thus, in any given sample, there is a certain percentage of neocytes and a certain percentage of gerocytes. Also, human blood contains a relatively large amount of iron, on the order of 108 mg/dl of red cells. Furthermore, the iron content is relatively uniform regardless of the average cell age of the blood sample. There are some patients, those suffering from chronic anemias for example, who depend upon repeated blood transfusions for their survival. Indeed, they may receive donor blood at such a rate that their systems are unable to entirely dispose of the iron content of that blood with the result that those patients suffer from iron overload and may die from complications resulting from this cause.
Since the contribution to iron overload is the same from the oldest transfused red cells which survive only a few hours as from the youngest ones which circulate in the body for months, it has been obvious for some time that a blood transfusion for patients such as this would be much more effective in terms of the ratio of physiological benefit to iron overload if the older red cells were removed from the donor blood and only the younger cells were administered to the patient.
It has also been recognized that the red cells in donor blood have a certain density distribution. Indeed, it turns out that the older red blood cells are more dense than the younger ones. Using this knowledge, attempts have been made to separate the red cells in a donor sample according to their densities so as to segregate the younger red cells or neocytes from the older cells or gerocytes. Some such attempts, described for example in the following publications, involve centrifuging the donor blood:
Murphy, John R., Influence of Temperature and Method of Centrifugation on the Separation of Erythrocytes, DJ. Lab. Clin. Med., August 1973, pp. 34-341; Corash, Lawrence M., et al, "Separation of Erythrocytes According to Age on a Simplified Density Gradient", J. Lab. Clin. Med., July, 1974, pp. 147-151; Piomelli, Sergio, et al, "Separation of Younger Red Cells With Improved Survival in Vivo: An Approach to Chronic Transfusion Therapy", Proc. Natl. Acad. Sci. USA 75 (1978), pp. 3474-3477; and Vettore, Luciano, et al, "A New Density Gradient System for the Separation of Human Red Blood Cells", American Journal of Hematology, 8:291 at Volume 8 (1980), pp. 291-297.
A centrifuge is usually used to separate different blood components by magnifying the different densities of the various blood components. Heretofore, the environment in which the blood bag is placed has been left to chance, with several factors having a negative influence on the quality of the separated cells. Typically, the blood bag filled with blood fluid is placed directly in a centrifuge cup along with 1, 2 or 3 empty satellite bags (used to receive the various separated components) connected to the blood bag by flexible plastic tubing, the entirety constituting an integral, fluid tight bag set. Rubber disks are used to balance the opposing centrifuge cups and are randomly placed upon or around the various bags in an uncontrolled manner. During centrifugation, the force exerted on the primary bag causes the blood fluid to compress into the bottom of the centrifuge cup. The manner in which the bag filled with blood fluid is compressed during centrifugation and the interaction of the associated empty bags upon compression with the filled bag are uncontrolled and left to chance.
At times, wrinkles or folds occur in the filled bag which trap heavier cells associated with the layer of the bag normally occupied by lighter cells, thus contaminating the various components with each other. In addition, the height-to-width ratio or aspect ratio of the blood fluid volume in the blood bag is random. The greater the ratio, the greater the distance cells must travel to reach their final density strata. The greater the distance, the larger the force and centrifugation time required to accomplish the separation. On the other hand, a maximum aspect ratio after centrifugation is advantageous because it minimizes the liklihood of inadvertant remixing of the separated cells.
At the end of centrifugation, the primary bag retains more or less the shape assumed during the centrifugation process. Thus, when the bag is compressed, the various separated components may be in close proximity (low aspect ratio) and subject to inadvertant remixing as the primary and satellite bags are pulled from the centrifuge cup. The tendency to remix may also be increased due to the primary and satellite bags wedging themselves together. The balancing disks further compound this problem.
After centrifugation, the first blood bag containing the blood components, separated into their density-specific components, is removed from the centrifuge cup and placed in an expressor which is a mechanical squeezing device. The tubes between the first bag and the empty satellite bags are either opened or pinched off according to the types of components to be transferred into those bags. The primary bag is then gently squeezed from bottom to top so that the upper layers of the blood volume are transferred to the satellite bags.
The removal of the bags from the centrifuge cup, their placement on the mechanical expressor and the monitoring of the correct volumes of the primary and satellite bags are steps which are time consuming, prone to technician error and may result in cross-contamination of the separated blood components.
One disadvantage of this known separation method is the occurance of contamination of one cell component with another due to the uncontrolled placement of the bags which may result in wrinkles that trap cells in the incorrect region of a bag. Another disadvantage of the known method is the time associated with the separation of the blood fluid into its density-dependent layers. If the unseparated blood column has a high aspect ratio, the cells of various densities have greater bag lengths to travel to reach their appropriate position. The greater the aspect ratio, the greater the spin force or time which must be used to accomplish the separation and the greater the spin force or time, the greater the cell component damage. This effect is particularly pronounced in the separation of platelets from PRP.
U.S. Pat. Nos. 4,416,778 and 4,582,606 disclose devices for harvesting neocytes. Both of these approaches involve removal of the older, more dense RBC (gerocytes) from the bottom of the RBC column after centrifugation. These approaches have merit, but will result in contamination of the young cells (neocytes) remaining in the primary bag due to adhesion of some of the older cells (gerocytes) to the primary bag walls. In addition, the older cells in both patented apparatus are transferred into round flexible bags. The percentage of RBC in the bags is often greater than 98% and the lack of plasma or other nutritional fluid in the bags may result in cell death. Further, the 98% RBC mass is not transfusable without the addition of solution to lower the RBC percentage to from 50% to 70%. The addition of such solution is not provided for in those patented apparatus, nor is there provision for withdrawing the RBCs to another bag for the dilution step or for any subsequent transfusion.
Still further, the entering of the lower bags of those prior devices through ports, which may be added to those bags, would result in breaking of those closed systems and, thus, require the RBCs to be used within 24 hours. These deficiencies can only be avoided by providing a bag containing the necessary nutritional fluid integrally attached to the bag containing the gerocytes. Such a solution does not appear to be feasible in either of those patented devices.